Process for preparing optically active 2-amino-ω-oxoalkanoic acid derivatives

ABSTRACT

A process is disclosed for preparing an (S)-2-amino-ω-oxoalkanoic acid derivative in which the corresponding aldehyde is converted into the corresponding acetal-protected aldehyde, the acetal-protected aldehyde is converted into the corresponding aminonitrile, the aminonitrile is converted into the corresponding amino acid amide, the amino acid amide is subjected to an enzymatic, enantioselective hydrolysis in which the (R)-enantiomer of the amino acid amide remains and the (S)-enantiomer is converted into the (S)-amino acid, and the (S)-amino acid is isolated. Preferably, the reaction mixture obtained after the conversion of the aminonitrile into the amino acid amide is treated with a benzaldehyde to form the Schiff base of the amino acid amide. The Schiff base is separated out and is converted into the free amino acid amide.

This is a division of application Ser. No. 09/160,342, filed Sep. 25,1998 now U.S. Pat. No. 6,133,002 which claims the benefit of ProvisionalApplication No. 60/069,776 filed Dec. 16, 1997.

RELATED APPLICATIONS

This application is a complete application that claims the benefit ofU.S. Provisional Application No. 60/069,776 filed Dec. 16, 1997, thecomplete disclosure of which is incorporated herein by reference.

FIELD OF THE INVENTION

The invention relates to a process for preparing an optically active 2amino-ω-oxoalkanoic acid derivative represented by formula 1

wherein n equals 0, 1, 2, 3, or 4 and R₁ and R₂ each independentlyrepresent an alkyl group with 1-10 carbon atoms or jointly form analkylene group to thereby form a ring with 3 or 4 carbon atoms togetherwith the oxygen atoms to which they are bound and the carbon atom towhich the oxygen atoms are bound. The starting materials for preparingformula 1 are readily commercially available.

BACKGROUND OF THE INVENTION

A preparation of a racemic mixture of a 2-amino-ω-oxoalkanoic acidderivative of formula 1 is described in Biorg. & Med. Chem. (1995)1237-1240. However, the preparation method described therein proceedsvia an 8-step process, starting from 3,4-dihydro-2H-pyran, with variousprotection and de-protection steps and is hence very laborious.

SUMMARY AND OBJECTS OF THE INVENTION

The invention provides a new, simple method for preparing2-amino-ω-oxoalkanoic acid derivatives represented by formula 1

wherein n equals 0, 1, 2, 3, or 4 and R₁ and R₂ each independentlyrepresent an alkyl group with 1-10 carbon atoms or jointly form analkylene group to thereby form a ring with 3 or 4 carbon atoms togetherwith the oxygen atoms to which they are bound and the carbon atom towhich the oxygen atoms are bound. The starting materials for preparing acompound of formula 1 are readily commercially available.

A process is disclosed for preparing an (S)-2-amino-ω-oxoalkanoic acidderivative in which the corresponding aldehyde is converted into thecorresponding acetal-protected aldehyde, the acetal-protected aldehydeis converted into the corresponding aminonitrile, the aminonitrile isconverted into the corresponding amino acid amide, the amino acid amideis subjected to an enzmatic, enantioselective hydrolysis in which the(R)-enantiomer of the amino acid amide remains and the (S)-enantiomer isconverted into the (S)-amino acid, and the (S)-amino acid is isolated.Preferably, the reaction mixture obtained after the conversion of theaminonitrile into the amino acid amide is treated with a benzaldehyde,in which the Schiff base of the amino acid amide is formed, the Schiffbase is separated and is converted into the free amino acid amide.

This is achieved according to the invention with a process convertingthe corresponding aldehyde of formula 2

wherein n is as described above into the corresponding acetal-protectedaldehyde of formula 3

wherein n, R₁ and R₂ are as described above,

converting the acetal-protected aldehyde into the correspondingaminonitrile of formula 4

wherein n, R₁ and R₂ are as described above,

converting the aminonitrile into the corresponding amino acid amide offormula 5

wherein n, R₁ and R₂ are as described above,

subjecting the amino acid amide to an enzymatic, enantioselectivehydrolysis in which the (R)-enantiomer of the amino acid amide remainsand the (S)-enantiomer is converted into the (S)-amino acid representedby formula (1), and

isolating the (S)amino acid.

It has been discovered that, in spite of the fact that the selectivityin the first step of the process can be relatively low, an economicallyattractive process can nevertheless be obtained.

The optically active compounds represented of formula 1 are novel andare particularly suitable for use in the preparation of, for example,allysin, an important crosslink precursor in proteins, as described inInt. J. Pept. Protein Res. (1988), 307-20, and in the preparation ofpharmaceuticals, for example as described in EP-A-629627, the completedisclosures of both are incorporated herein by reference.

Allysin can be obtained by an acid catalysed (e.g. an Amberlist -15)deacetylation of a compound of formula 1 with n=3.

Without protection such compound will be immediately converted in thecyclic compound:

For use in the peptide synthesis a compound of formula 1 with n=3 istreated with HCl and MeOH in order to obtain

An example of peptide chemistry is shown in FIG. 1, using the abovecompound (8), and is also further described in J. Pept. Protein Res.(1988), 307-20, incorporated by reference above.

The preparation of a pharmaceutical as referred to above is shown inFIG. 2, and is described in EP-A-629627, incorporated by referenceabove.

More particularly, in the process according to the invention, one of thetwo aldehyde functional groups in the aldehyde represented by formula 2is first protected through conversion, in a manner known per se, into anacetal. cf. T. H. Greene, Protective Groups in Organic Synthesis, JohnWiley & Sons, New York (1981). This can for example be done with the aidof an alcohol, for example an alcohol with 1-5 carbon atoms when R₁ andR₂ represent an alkyl group, or with the aid of a diol with 1 to 5carbon atoms, in particular a 1,2-ethanediol or a 1,3-propanediol,whether or not substituted with for example an alkyl group with 1-5carbon atoms, for example 1,2-ethanediol, 1,2-propanediol,1,3-propanediol or 2,3-butanediol, when R₁ and R₂ form part of a ringstructure; or via re-acetalization, for example with the aid ofortho-formate esters.

The acetalization can, for example, be carried out by bringing thealdehyde represented by formula 2 into contact with an alcohol or a diolunder acid conditions such as, for example, in the presence of asulphonic acid, in particular p-toluenesulphonic acid. The acetalizationis optionally carried out in the presence of a solvent. In principle,any solvent that does not interfere with the reaction can be used as asolvent such as, for example, aromatic hydrocarbons, in particularbenzene, toluene and xylene; halogenated hydrocarbons, for exampledichloromethane; esters, preferably hindered esters, in particularisopropyl acetate or isobutyl acetate or other esters having bulky estergroups; and ethers, in particular methyl-t-butyl ether (MTBE). Theacetalization with an alcohol or diol is preferably carried out atelevated temperature, for example at a temperature of between about 50°C. and about 150° C., preferably at reflux temperature.

The acetal-protected aldehyde obtained is subsequently converted intothe corresponding aminonitrile, for example via a Strecker synthesis.cf., J. March, Advanced Organic Chemistry, John Wiley & Sons, New York,pp. 855-856 (3^(rd) Ed. 1981); T. W. Graham Solomons, Fundamentals ofOrganic Chemistry, John Wiley & Sons, New York, pp. 980-981 (4^(th) Ed.1994). The acetal-protected aldehyde can, for example, be converted intothe amnonitrile in the presence of ammonia with the aid of a cyanidecompound, for example HCN, NaCN or KCN.

The aminonitrile compounds represented by formula 4 intermediates arenovel compounds. The invention hence also relates to theseintermediates. The racemic (R,S)-amide mixture is enantioselectivelyhydrolysed using an enzyme, so as to give the (R)-amide and the(S)-acid. In order to obtain the (R)-acid the (R)-amide is subsequentlyhydrolysed into the (R)-acid.

The aminonitrile represented by formula 4 is subsequently converted intothe corresponding racemic amino acid amine. One suitable method isdescribed, for instance, in GB-A-1548032, the disclosure of which isherein incorporated by reference. The aminonitrile is converted at a pHof between 11 and 14, preferably between 12.5 and 13.5, in the presenceof a base and a ketone or aldehyde, optionally followed by hydrolysis ofthe intermediately formed Schiff base in the presence of water.Preferably, an alkali metal hydroxide, such as KOH or NaOH, or acorresponding base is used as the base and an aliphatic ketone, forexample acetone, methyl ethyl ketone or cyclohexanone, or an aromaticaldehyde, for example benzaldehyde, is used as the ketone or aldehyde.

Aminonitile obtained as formula 4 can be isolated via extraction fromthe aqueous phase using an organic solvent (e.g. aromatic hydrocarbonssuch as toluene, esters or chlorinated hydrocarbons), and separation ofthe two layers, if desired. The organic layer containing theaminonitrile may be dried to give the free amino acid nitrile or treatedwith e.g. HCl-gas in order to precipate the (more stable) aminonitrileHCl salt.

A benzaldehyde is preferably used in the formation of the Schiff base ofthe amino acid amide. An advantage of benzaldehyde is that it is easy toseparate the Schiff base and recover the benzaldehyde. The Schiff baseof the amide is obtained as a precipitate and can be filtered off (andoptionally converted into the free amide). Another advantage ofbenzaldehyde is that it is not miscible with water, and consequently itis also far more preferable than other extraction means because the thusformed Schiff base of the optically active amino acid amide dissolves inthe benzaldehyde whereas the other components of the reaction mixtureare in the water phase. It has surprisingly been found that thehydrolysis of the Schiff base of the amino acid amide to obtain the saltof the amino acid amide can be carried out without the acetal functiondeteriorating significantly.

‘Benzaldehyde’ is also understood to include substituted benzaldehydessuch as lower (1-4 carbon atoms) alkylbenzaldehydes,halogenbenzaldehydes, nitrobenzaldehydes and lower (1-4 carbon atoms)alkoxybenzaldehydes.

The reaction with benzaldehyde resulting in the formation of a Schiffbase can, for example, be carried at a temperature of between about 20°C. and about 60° C., preferably between about 35° C. and about 45° C. Aprecipitate of the Schiff base of the amino acid amide is obtainedequimolar amounts of benzaldehyde, for example 0.9-2, in particular0.95-1.1 equivalents relative to the amino acid amide, are used in theformation of the Schiff base without a different solvent for the Schiffbase of the amide. The other components remain dissolved in the motherliquor. If an excess of benzaldehyde is used, the benzaldehyde acts bothas a reaction means and as a solvent, and two layers are obtained. It isalso possible to use mixtures of benzaldehyde and other solvents, forexample mixtures with aromatic hydrocarbons, for example toluene,ketones, for example methyl isobutyl ketone, halogenated hydrocarbons,for example chloroform or dichloromethane; esters, for example ethylacetate and butyl acetate. The organic phase can subsequently be used assuch in the hydrolysis of the Schiff base of the amino acid amide to theamino acid amide or it can be subjected to concentration, upon which theSchiff base of the amino acid amide precipitates as a solid.

The amino acid amide can be recovered from the corresponding Schiff basein a simple manner, through acidification with a strong acid, forexample sulphuric acid, until a pH of between 3 and 5, preferablybetween 3.5 and 4.5 has been obtained, with the Schiff base decomposingto form the aldehyde and the corresponding salt of the amino acid amide.

The free amino acid amide can subsequently be obtained from the saltthrough treatment with a base, for example with triethylamine.Preferably, the conversion of the salt into the free amino acid amide iscarried out with the aid of a (strongly) basic ion exchanger, forexample Amberlyst 26 or IRA 900.

The amino acid amide intermediates represented by formula 5 and theSchiff bases thereof that are represented by formula 6 are novelcompounds.

In the formula 6, n is 0, 1, 2, 3 or 4; R₁ and R₂ each independentlyrepresent an alkyl group having 1-10 carbon atoms, or R₁ and R₂ form anakylene group whereby a ring is formed; and from 1 to 4 R^(i)substituents are present, each R^(i) being independently selected fromamong halogen, nitro, an alkyl group or an alkoxy group. Representativealkyl groups having 1-10 carbon atoms include, among others, methyl,ethyl n-propyl, butyl (n-butyl, sec-butyl, t-butyl), pentyl andcyclopentyl. Halogen substituents are, independent of one another,fluoro, chloro, bromo and, in principle, iodo. The R^(i) groups areindependent of one another. The above-mentioned ring is formed with 3-4carbon atoms together with the oxygen atoms to which they are bound andthe carbon atom to which the oxygen atoms are bound. The alkyl andalkoxy group for R^(i) preferably contain 1-4 carbon atoms. Theinvention therefore also relates to these intermediates, both in racemicform and in optically active form, in particular the amino acid amideshaving an enantiomeric excess (“e.e.”) greater than 80%, preferablygreater than 90%, more preferably greater than 95%, most preferablygreater than 98%, in particular greater than 99%.

The racemic (R,S) amino acid amide is subsequently subjected to anenantioselective, enzymatic hydrolysis in which the (S)-enantiomer isselectively converted into the corresponding (S)-acid and the (R)-amideenantiomer remains unaffected. The (R)-amide may subsequently behydrolyzed into the (R)-acid. Also, the (S)-amide could be obtainedusing a D-amidase, as described in Yasuhisa Asano et al. Biochemical andBiophysical Research Communications, Vol 162, No. 1, 1989, p 470-474, oralternatively, via esterification and amidation of the (S)-amino acid.The (S)-2-amino-ω-oxoalkanoic acid derivative can then be obtained withan e.e. of more than 90%, in particular more than 95%, preferably morethan 98%, in particular more than 99%. The enantioselective enzymatichydrolysis is preferably carried out in an aqueous environment. It ishowever also possible to use an organic solvent. The temperature is notparticularly critical and lies, for example, between about 0° C. andabout 60° C., preferably between about 20° C. and about 50° C. The pH atwhich the enzymatic hydrolysis is carried out is preferably between 5and 10.5, in particular between 8.0 and 9.5. Suitable, amidase enzymesfor example, are available, and include, an amidase derived from thegenus Asperillus, Mycobacterium, Aeromonas, Bacillus, Pseudomonas orOchrobactrum. Preferably, an amidase derived from Pseudomonas putida orfrom Ochrobactrum anthropi is used.

After, for example, the removal of the (R)-enantiomer of the amino acidamide as the Schiff base, or concentration and treatment with alcohol,for example isopropanol, the optically active (S)-2-amino-ω-oxoalkanoicacid derivative can be obtained. The Schiff base of the (R)-enantiomerof the amino acid amide can, optically, be converted into the free aminoacid amide, which in turn can optionally be hydrolyzed under mildconditions, for example via a (non-stereoselective) enzymatic hydrolysisas described in EP-A-179523, the disclosure of which is hereinincorporated by reference, using Rhodococcus erythropolis or an extractthereof, to obtain the (R)-2-amino-ω-oxoalkanoic acid derivative. Theoptically active (R)-2-amino-ω-oxoalkanoic acid derivatives are novelcompounds with various applications. The compounds represented byformula 7

are, for example, particularly suitable for use in the preparation ofD-pipecolic acid derivatives such as, for example, described in J.O.C.(1990), 5551-3, and in Bioorg. & Med. Chem. (1995), 1237-1240, thedisclosures of which are both herein incorporated by reference, whichare in turn used per se in the preparation of various pharmaceuticalssuch as, for example, described in EP-A-672665, DE-A-3702943 and U.S.Pat. No. 5,409,946, the disclosures of all three being hereinincorporated by reference. The compounds represented by formula 7 canalso be used as an intermediate(s) in the preparation of D-proline,which is used, for example, in the preparation of Elitriptan asdescribed in ‘Drugs of the Future’ (1997), 221-223, the disclosure ofwhich is herein incorporated by reference. The invention also relates tothese novel compounds, in particular to the optically active(R)-2-amino-ω-oxoalkanoic acid derivatives having an e.e. greater than80%, preferably greater than 90%, in particular greater than 95%.

The preparation of D-pipecolic acid and D-proline, referred to above inprinciple analogous. Reaction schemes are shown in FIG. 3. The subjectmatter of B. Ohtani et al., J. Org. Chem. (1990), 55, 5551-5553, and T.F. Buckley and H. Rapaport, J. Am. Chem. Soc. (1982) 104, 4446-4450,referred to in FIG. 3 is also incorporated herein by reference.

An application of these compounds is given in FIG. 3. According to FIG.3-2 and FIG. 3-3, compound 9 (obtained from the formula 1 compound withn=3) is converted in 10, 11, 12, and 3 subsequently and compound 3 is,together with compound 2 converted into compound 1, shown in FIG. 3-2.

The use of D-proline is demonstrated in FIG. 3-3 wherein compound 14represents D-proline.

The description of the invention in Netherlands Application 1007113,filed Sep. 25, 1997 is incorporated herein by reference.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a reaction scheme using compound (8) in the synthesis.

FIG. 2 is a reaction scheme taken from EP-A-629627.

FIG. 3 is a reaction scheme using D-proline.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

The invention will now be elucidated with reference to the exampleswithout being limited thereby.

EXAMPLES Example I Preparation of 4-(1.3-dioxolane-2-yl)-1-butanal viathe acetalization of glutardialdehyde with ethylene glycol

Toluene (1 liter), ethylene glycol (155 grams, 2.5 mol) andp-toluenesulphonic acid (1 gram) were successively dosed to a 50%solution of glutardialdehyde in water (500 grams, 2.5 mol). The mixturewas heated to reflux temperature. The water was azeotropically removedwith the aid of a Dean-Stark apparatus. As soon as all the water hadbeen removed with the aid of the Dean-Stark apparatus (approx. 6 hours)the solution was cooled to room temperature.

Sodium bicarbonate (2.1 grams) and water (250 ml) were added to thesolution. After vigorous stirring for half an hour the layers wereseparated. This washing of the toluene phase was repeated twice. Thenthe toluene phase was evaporated in a rotary film evaporator to obtain apale yellow oil. The 4-(1,3-dioxolane-2-yl)-1-butanal content was 50%(G.C.) with a yield 5 of 35%.

Example II Preparation of the Schiff base of2-amino-5-(1,3-dioxolane-2-yl)-pentanoic acid amide via the Streckerreaction of 4-(1,3-dioxolan-2-yl)-1-butanal

Sodium cyanide (49 grams, 1 mol) and ammonium acetate (77 grams, 1 mol)were successively added to a 25% solution of ammonia in water (500 ml).The oil obtained in Example I (288 grams, 1 mol) was added dropwise tothis solution over a period of one hour. After further stirring for 5hours, acetone (100 ml) and a 45% solution of potassium hydroxide inwater (10 ml) were successively dosed to this solution. After 3 hoursglacial acetic acid (7 ml) was added. The solution was concentrated toapprox. 400 grams with the aid of a rotary film evaporator to obtain aconcentrate. Water (600 ml) and toluene (150 ml) were added to theconcentrate. After stirring for half an hour layers were separated.Benzaldehyde (74 grams) was slowly dosed to the water phase withvigorous stirring. The white precipitate formed was removed throughfiltration and washed with water. After drying, the yield of the Schiffbase of 2-amino-5-(1,3-dioxolane-2-yl)-pentanoic acid amide was 195grams (yield 70%) at a purity >98% (¹H-NMR).

Example III Preparation of (S)2-amino-5-(1,3-dioxolane-2-yl)pentanoicacid via the enzymatic resolution on2-amino-5-(1,3-dioxolane-2-yl)-pentanoic acid amide

The Schiff base of 2-amino-5-(1,3-dioxolane-2-yl)-pentanoic acid amide(240 grams, 0.87 mol) was suspended in toluene (1000 ml) and water (1500ml). Concentrated sulphuric acid (44.5 grams, 0.44 mol) was very slowlydosed with vigorous stirring, so that the pH remained above 4.0. Afterthe dosage of the sulphuric acid the layers were separated. The waterphase was subsequently passed over a strongly basic ion exchanger (typeIRA 900). The sulphate-free water phase was subsequently brought to a pHof 9.0 with acetic acid. Pseudomonas putida (obtained from NOVO) (40grams) was added to this solution. After stirring at 37° C. for 8 hours,decalite (30 grams) and, dropwise, benzaldehyde (51 grams, 0.48 mol)were successively dosed to the suspension. The precipitate formed wasremoved with the aid of filtration. The filtrate obtained wasconcentrated to about 450 grams with the aid of a rotary filmevaporator. After heating to 65° C. isopropanol (900 grams) was added tothe solution. After slow cooling to −5° C. the white crystals formedwere filtered and successively washed with ice water and isopropanol.The yield of (S)-2-amino-5-(1,3-dioxolane-2-yl)-pentanoic acid was 42grams (26%). The melting point was 258° C., the purity was >99%(titration), and the e.e % was greater than 99% (HPLC).

Example IV Preparation of (S)-2-amino-5-(1,3-dioxolane-2-yl)-pentanoicacid via the enzymatic resolution on2-amino-5-(1,3-dioxolane-2-yl)-pentanoic acid amide

Glacial acetic acid was added to the water phase obtained after thetoluene extraction (see Example II) so that the pH became 9.0. At 37° C.Pseudomonas putida (obtained from NOVO) was added to this water phase.After stiring for 8 hours, the water phase was treated as described inExample III. The yield of (S)-2-amino-5-(1,3-dioxolane-2-yl)-pentanoicacid was 11%, the purity was 95% (HPLC), and the e.e. % was >99% (HPLC).

Example V Preparation of (S)-2-amino-5-(1,3-dioxolane-2-yl)pentanoicacid via the enzymatic resolution on2-amino5-(1.3-dioxolane-2-yl)-pentanoic acid amide

The water phase obtained after the hydrolysis of the Schiff base withsulphuric acid (see Example III) was brought to a pH of 9.0 with the aidof triethylamine. At 37° C., Pseudomonas putida (obtained from NOVO) wassubsequently dosed to this solution. After stirring for 8 hours, thethus obtained suspension was processed further as described in ExampleIII. The yields of (S)-2-amino-5-(1,3-dioxolane-2-yl)-pentanoic acid was24%. The purity was 76% (HPLC) and the e.e. % was greater than 99%(HPLC).

What we claim is:
 1. The 2-amino acid amide represented by the formula 5

or the Schiff base thereof represented by the formula 6,

wherein n equals 0, 1, 2, 3 or 4; R₁ and R₂ each independently representan alkyl group with 1-10 carbon atoms or form a ring with 3 or 4 carbonatoms together with the oxygen atoms to which they are bound and thecarbon atom to which the oxygen atoms are bound; and 1-4 R^(i) groups,each of which is independently selected from the group consisting ofhalogen, nitro, an alkyl group and alkoxy group with 1-4 carbon atoms.2. An optically active compound according to claim 1 wherein theoptically active compound has an e.e. of >95%.
 3. An optically activecompound according to claim 2, wherein the e.e. is greater than 98%. 4.The 2-amino acid amide according to claim 1 wherein n is 2 or
 3. 5. Acompound according to claim 1, wherein R₁ and R₂ form a ring with 3 or 4carbon atoms together with the oxygen atoms to which they are bound andthe carbon atom to which the oxygen atoms are bound.
 6. A Schiff base offormula 6 formed from benzaldehyde and the 2-amino acid amide of formula5 of claim
 1. 7. A Schiff base according to claim 6 obtained as aprecipitate.